Abstract

Myotonic dystrophy type 1 (DM1) is a multisystem disease and the most common adult form of muscular dystrophy, leading to severe, progressive muscle wasting and weakness, but which also impacts the eye, heart and brain. It can also affect infants and results in severe disability and significantly shortened life span. No treatment is presently available. The disease is caused by a genetic defect in the DMPK gene, which when mutated results in the formation of a toxic product that directly leads to disease.

We have developed a novel method based on short synthetic DNA drugs (known as antisense oligonucleotides) which bind to and inactivate the mutated gene product, and which we attach to potent protein fragments or peptides which allows the DNA drug to be delivered with very high efficiency to ALL of the affected tissues including muscle, heart and brain. We have already demonstrated the proof that this approach can work both in cell cultures and also in DM1 animal models. However, there is still room for improvement in the delivery of the treatment to muscle (especially diaphragm) and heart which is especially critical since the life span of DM1 patients is reduced particularly due to respiratory insufficiency and cardiac failure. Here we now plan to capitalise on our peptide technology to select and optimise the best delivery peptide which would permit highly effective delivery of the DNA drug to all affected tissues in the absence of unwanted side effects. Our laboratories are world-leading and have all necessary skills to deliver the project plan.

The major objective of this proposal is to develop this peptide-DNA method into a drug that could be used in patients. More specifically the objectives are to (1) screen a number of peptide-DNA compounds for their ability to treat the main features of the disease in human cells; (2) optimise lead and back-up compounds selected for high efficiency activity in the tissues most affected in a DM1 mouse model, in the absence of unwanted side effects; (3) establish the doses at which the treatment is most effective and how long the effects last.

This is a highly innovative project that meets an unmet clinical need and could have a very high clinical impact but we first need to optimize the delivery of the treatment to the tissues most affected by DM1 (heart and diaphragm) and that represent the major limiting factor for DM1 patient survival, before we start clinical development of the compounds for evaluation in patients. To date the only treatments for DM1 target some of the symptoms such as the removal of cataracts or hormonal replacement therapy but there are no treatments that directly target the cause of the disease. Therefore, the optimization of these peptide-DNA compounds able to block the product of the mutated gene is a high priority for preclinical and clinical development. Additionally, an anticipated output of this project is intellectual property involving the most successful compounds. Moreover, the improvement of compound delivery into muscles would be a major step in the development of an effective therapy not only for DM1 but potentially for other muscular dystrophies.

Technical Summary

Myotonic dystrophy type 1 (DM1) is the most common adult form of muscular dystrophy. A severe congenital form affects infants and children resulting in severe disability and shortened life span. No treatment is presently available for DM1. The genetic basis of DM1 is a CTG repeat expansion in the 3'UTR of the DMPK gene. The favoured pathogenic mechanism involves RNA gain-of-function whereby repeat-containing transcripts from the expanded allele accumulate in affected cell nuclei where they sequester nuclear proteins involved alternative splicing regulation. This leads to dysregulated pre-mRNA splicing resulting in a multisystem disorder, affecting muscle, heart and brain.

Antisense oligonucleotides (AO) are single-stranded nucleic acids which, when directed toward the repeat expansion, prevent sequestration of nuclear proteins to the CUG repeats. Proof-of-principle has already been demonstrated for this approach, however current generation AOs show poor distribution to muscle, including diaphragm and heart, and do not penetrate the blood brain barrier. This is critically important for disease modification since DM1 life span in is reduced due to respiratory and cardiac failure.

Our SOLUTION is an advanced peptide delivery technology allowing development of peptide conjugated AOs of morpholino (PMO) chemistry thereby permitting highly effective systemic peptide-PMO delivery to all DM1 affected tissues. Starting with well-established (CAG)n PMOs, we will select and optimise peptide sequence, building on the well-established Pip peptide series, to identify lead and back-up peptide-PMOs with high efficacy both in vitro and in vivo. The most effective peptide-PMOs will be selected on the basis of efficacy and safety to determine an optimal treatment dose, including multi-dose, long-term treatment regimens, in model DM1 mice. This project will therefore identify an optimised, lead peptide-PMO compound suitable for clinical development and evaluation in DM1 patients.

Planned Impact

We anticipate that our research programme on the development of an advanced antisense oligonucleotide (AO) therapy for Myotonic Dystrophy type 1 (DM1) patients will have a range of major impacts including academic, health, socioeconomic and commercial.

A range of likely academic beneficiaries have already been identified and it is likely that the proposed work, if successful, will have a high ACADEMIC IMPACT on wide range of fundamental scientific and clinical academic groups.

Naturally the most important impact would be the HEALTH IMPACT on the future health outcomes for DM1 patients, their families and associated advocacy groups. If successful it is likely that this work could lead to development of an AO based disease modifying therapy for evaluation in DM1 patients within 5 years. If successful, it is also likely that this research programme will lead to significant impacts on related neuromuscular diseases and hence it is possible that large numbers of patients will ultimately benefit from this novel platform peptide technology. Although we have not yet undertaken a health economic analysis of the likely benefits arising from successful development of a peptide-AO therapy for DM1, it seems highly likely that successful development of such a disease modifying therapy is likely to result in SOCIOECONOMIC IMPACT via reduced healthcare costs, reduced hospital stays and reduced need for advanced medical interventions.

In addition to these academic, health and socioeconomic impacts outlined above, this preclinical development programme could have a major COMMERCIAL IMPACT on the biotech / pharma industry, which has already expressed strong interest in the potential of advanced antisense oligonucleotide methods for treating neuromuscular diseases.

There is an increasing number of biotech / pharma companies currently developing or with strong interest in developing antisense oligonucleotide therapies (BioMarin, Ionis Pharmaceuticals, Pfizer, Santaris, Sarepta Therapeutics, Shire etc). The reason why being that antisense oligonucleotides have shown promise to target the RNA of genes involved in disease that cannot be treated efficiently with any other method. However, the difficulty of delivering the oligonucleotide treatment to the tissues most affected by a number of pathologies as is the case for neuromuscular disease has thus far greatly limited the success in the clinic since the systemic delivery of antisense oligonucleotides tends to lead to non-specific accumulation of the treatment in a few peripheral organs, mainly in the liver. In fact, the only recent FDA approval of an antisense compound is Kynamro which is an oligonucleotide inhibitor of apolipoprotein B-100 synthesis aiming to reduce cholesterol levels by preventing the synthesis in the liver of atherogenic lipoproteins. Therefore, there is a real need in industry and improved delivery is the key area to develop effective oligonucleotide therapies likely to gain FDA approval. Our proposed programme does just that by conjugating the antisense molecules to peptides that allow them to have effective body-wide biodistribution and, thus, our technology has the potential to have a high impact on the pharmacological industry.

Although we still do not have an industrial partner to commercialize our DM1 therapy we have already established strong links with industry over the last few years, working with Pfizer, Shire and Sarepta Therapeutics in our DMD disease programme. We will continue to work closely with Oxford University Innovation, the technology transfer company of the University of Oxford, that will manage all intellectual property developed during the course of the programme and protect the IP generated before engaging with an industrial partner. This strategy will help us to ensure that the proposed programme has the desired commercial and health impact on area of high unmet medical need.

Preclinical development of antisense oligonucleotides for Myotonic Dystrophy 1.

Organisation

Sorbonne Universités

PI Contribution

Peptide and oligonucleotide design, synthesis, and quality control of peptide-oligonucleotides is performed in the Wood Laboratory. Our laboratory has access to a purpose-built peptide synthesis facility with state of the art equipment and provides a very high quality intellectual and physical environment for peptide and nucleic acid chemistry. The Wood laboratory also performs the analysis of sequencing data and biodistribution of the compounds.

Collaborator Contribution

The team of Denis Furling are specialists in pathology of Myotonic Dystrophy 1. They have a well stablished HSA-LR mouse colony that recapitulates the main aspects of the disease and have developed a number of immortalized cells lines derived from patients. The Furling laboratory has also optimized a technique to measure the myotonia in vivo.